Field of the Invention
The present invention relates to a method for stabilising fatty acids present in a sample such as bodily fluids. The present invention further relates to a solid medium which is capable of stabilising fatty acids applied thereto, and a method for preparing same. The present invention further relates to a method for determining the fatty acid composition of a sample.
Description of Related Art
A high intake of omega-3 fatty acids especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) has been associated with reductions in cardiovascular disease, neurodegenerative diseases, diabetes and arthritis in adults and improved mental development in preterm infants. As a result, health authorities around the world are now recommending that intake of omega-3 fatty acids be increased, for example by consuming 2 to 3 meals including fish per week.
Dietary omega-3 fatty acid recommendations for cardiac protection include increasing fish intake to two fatty fish meals per week or consuming 600 or 400 mg/day long chain omega-3 fatty acids respectively for men and women. Although it is known that these recommendations will increase cellular omega-3 fatty acid levels to a level within the cardioprotective range in clinical trials, there is almost no knowledge about how these recommendations are influencing the fatty acid status of the general population. Because the fatty acid status in blood is a useful marker for evaluating cardiac risk in human populations and identifying those individuals who would benefit from dietary omega-3 fatty acid supplementation, evaluation of fatty acid status is important.
Fatty acid profiling of human blood samples which are used as a marker of fatty acid status in human organs has become an important tool for understanding the relationship between dietary fatty acid intake and fatty acid status, since blood levels are thought to reflect biological actions and blood is accessible for collection in human studies. However, conventional approaches to assaying fatty acids in blood involve venous blood collection and an expensive, time consuming multi-step processes that limits its usefulness as a screening tool. Accordingly, a rapid, cheap and reliable means of measuring cellular omega-3 fatty acid levels is necessary.
Until recently, such a rapid and reliable test for the level of omega-3 fatty acids present in human blood (known as The Omega 3 Index) has not been available. In the past few years several companies have advertised a blood spot test in which, following the pricking of a finger, a blood spot is placed on a small piece of filter paper with or without antioxidant(s). After drying, the filter paper is sent to a central laboratory for analysis. It has been claimed that the fatty acids in the blood spot are stable for weeks and that a reliable measure of the Omega 3 Index can be obtained. The present inventors have made attempts to replicate these tests and found that significant degradation of fatty acids occurs over time.
One such product is the Fluka blood collection kit developed by Sigma-Aldrich for the direct assessment of n-3 and n-6 long-chain polyunsaturated fatty acids in blood spots. The only long term stability study to be conducted on blood spot samples collected on Fluka kit papers stored at room temperature showed that there was a significant decline in the long-chain polyunsaturated fatty acid levels (as a percentage of total fatty acids) after 1 month (Min et al. Prostaglandins, Leukotrienes and Essential Fatty Acids, 2011 84, 13-18). Thus, the oxidation of long-chain polyunsaturated fatty acids makes this blood spot method impractical for use since it would necessitate that all samples were analysed at a fixed time after collection if results were to be compared.
There is therefore a need for a simple, rapid and reliable method for determining fatty acid content in blood wherein fatty acid degradation during storage of the blood is minimised or eliminated.
Surprisingly, the present inventors have discovered that fatty acids within a dried blood spot may be stabilised for a period of months at ambient conditions. The present inventors have further discovered that a highly accurate determination of the fatty acid composition of blood stored on a solid medium for a period of months may be achieved by stabilising the fatty acids and by controlling contaminants in the solid medium.